Feasibility of a recombinant human apolipoprotein E reference material
| Autor: | Françoise Schiele, L Havekes, Lawrence P. Aggerbeck, Maryvonne Rosseneu, Athanase Visvikis, C Profilis, Gérard Siest, Ann Barbier, M Huttinger |
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| Přispěvatelé: | Gaubius Instituut TNO |
| Jazyk: | angličtina |
| Rok vydání: | 1998 |
| Předmět: | |
| Zdroj: | Scopus-Elsevier Fresenius' Journal of Analytical Chemistry, 3-4, 360, 501-504 |
| Popis: | The aim of this work was to prepare a recombinant apo E material and to determine its suitability as a reference material. We produced human apo E3 using recombinant DNA technology. The cDNA of human apo E3 was cloned in the pARHS bacterial expression vector and used to transfect E. Coli BL21 (DE3) cells. The recombinant protein was then purified in one step by affinity chromatography on a Ni-chelated agarose column under denaturing conditions. The purity of the protein estimated by SDS PAGE was greater than 96%. The physicochemical properties and biological and immunological reactivity of the purified recombinant apo E3 were shown to be close to those of the protein purified from human plasma VLDL. A limited batch of lyophilized apo E material was then prepared. The stability of the lyophilized apo E material examined by temperature accelerated degradation was acceptable. No degradation of the measured apo E was observed after storage of the lyophilized material at +4°C and -20°C for 11 months. The reconstituted lyophilized material, in comparison with human fresh serum samples and with apo E purified from human VLDL, showed no major alteration of its immunological reactivity when assayed by immunoturbidimetry or ELISA. © Springer-Verlag 1998. |
| Databáze: | OpenAIRE |
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